The invention relates to novel Agaricus bisporus hybrids, a method of obtaining them, and their use in mushroom growing,.
It is known that growing the mushroom Agaricus bisporus, also called cultivated mushroom, comprises a certain number of steps, which are recalled herein-below.
A first step (composting) consists in the preparation of a substrate based on straw, horse manure and, if appropriate, nitrogen-containing additives, in order to obtain, by fermentation, a substrate (compost) which contains nutrient elements which can be used by the mycelium during the multiplication phase. The duration of this composting step is around 2 to 3 weeks. In general, the composting step is completed by a step called pasteurization, which consists in improving the quality of the compost by directed fermentation in special rooms, in general over 5 to 7 days. The following step is inoculation, which consists in seeding the culture substrate as obtained with xe2x80x9cspawnxe2x80x9d, that is to say mycelium multiplied on cereal grains. The inoculation rate specifies the ratio of spawn weight to compost weight. This inoculation rate is generally over 0.5% and under 1%. The next step is the incubation, during which the mycelium multiplies and progressively colonizes the culture substrate. This incubation phase is carried out either on special controlled-environment premises or in a culture cave system at a substrate temperature of around 24-26xc2x0. The average time of the incubation phase is approximately 14 days. The next operation which is carried out is called casing, which consists in covering the culture substrate with a layer of a casing material which generally, in particular in France, consists of a mixture of limestone (for example tufa) and peat. This layer has a thickness of generally around 3 to 5 cm. To enhance the formation of fruiting bodies, a thermal shock technique is used which consists in lowering the ambient temperature to around 16-17xc2x0 C. with a substantial renewal of air and regular watering. This latter phase of inducing the formation of fruiting bodies is sometimes preceded, just after casing, by a phase called prefructification, which consists in maintaining an ambient temperature of around 24xc2x0 C. with slight renewal of air over 8 to 10 days.
The mushrooms are harvested from successive emergences (termed xe2x80x9cflushesxe2x80x9d). The so-called first-flush harvest can generally be carried out three weeks after casing. In general, it is possible to harvest 4 to 5 flushes over a period of 5 to 7 weeks after the beginning of the first harvest. The total harvest yield can be expressed by weight of mushrooms per 100 kg of compost. The average yields are approximately 30%.
Thus, growing cultivated mushrooms is very restricting, in particular concerning the temperature regulation, which leads to a noteworthy consumption of energy and thus to considerable costs.
The casing step is also expensive in terms of labour and casing material.
If the formation of the cultured mushroom is to be achieved, however, the casing operation cannot be dispensed with. The same applies to the thermal shock step.
The following works on methods of growing cultivated mushrooms can be cited in particular: J. Delmas, Les champignons et leur culture [Mushrooms and their production], published by La Maison Rustiquexe2x80x94Flammarion (Paris, France); The cultivation of Mushrooms (1988), published by Somycel; and Olivier J. M. et al., La culture des champignons [Mushroom production] (1991), published by Armand Colin. With reference to the casing step and the fact that it is required for obtaining the formation of fruiting bodies, W. A. Hayes, The Casing Layer, published by W. S. Maney and Son Ltd., Leeds (Great Britain) may also be cited.
The materials used for casing are very varied. Apart from peat and mixtures of peat and limestone rock, there may be mentioned clay, gypsum, sand and the like. The use of layers of synthetic materials or natural or synthetic polymeric materials which are capable of retaining water has also been recommended. In this matter, the patents, or patent applications, FR-2 549 346, FR-2 603 767, FR-2 676 158, FR-2 723 506 and U.S. Pat. No. 4,079,543 may be cited in particular.
It has now been found that it is possible to obtain Agaricus hybrids which can form fruiting bodies without casing and which can be grown under conditions which are less constraining as regards temperature regulation, because it has been found that certain hybrids obtained by hybridizing strains of Agaricus bisporus with Agaricus bisporus var. burnettii, or progeny or hybridization products of this variety, are capable of forming fruiting bodies without casing and can, moreover, form fruiting bodies without thermal shock. Moreover, the invention allows hybrids to be obtained whose mycelia are capable of multiplying on substrates at higher temperatures without losing their capability of forming fruiting bodies.
These novel hybrids can thus be grown while substantially reducing labour and energy costs.
Moreover, the invention makes it possible to grow cultivated mushrooms even in countries with a hot climate, where it was hitherto impossible to grow them profitably.
The invention relates to a hybrid mycelium of Agaricus bisporus having at least one of the following phenotypic, stable and genetically transmissible traits:
after multiplication on a usual culture substrate and penetration of this substrate, it is capable of forming fruiting bodies without casing;
after multiplication on a usual culture substrate and penetration of this substrate, it is capable of forming fruiting bodies at a substrate temperature of 26xc2x0 C. or above;
after multiplication on a usual culture substrate and penetration of this substrate, it is capable of forming fruiting bodies at an ambient temperature of 24xc2x0 C. or above;
it is capable of growing on a usual culture substrate at a substrate temperature of 30xc2x0 C. or above.
The term xe2x80x9chybrid myceliumxe2x80x9d designates in the present application:
a) either a mycelium which originates from at least one hybridization between two parental strains of Agaricus bisporus, that is to say which originates from the combination via fusion of cytoplasms (plasmogamy) of two homokaryons which themselves originate from two distinct heterokaryotic parent strains,
b) or a mycelium which originates from a spore produced by the fruiting bodies of a mycelium as defined under a) above,
c) or a mycelium which originates from a protoplast obtained from a mycelium as defined under a) and/or b) above.
Mycelia according to the invention are thus intraspecific hybrids.
When the hybrid mycelium according to the invention is capable of growing on a usual culture substrate at a temperature of, for example, 30xc2x0 C. or above, it must be understood that it is capable of penetrating this substrate at this temperature, without, however, losing its capability of forming fruiting bodies, which means, in particular, that fruiting bodies are produced within normal time limits (not more than 5 to 6 weeks after inoculation), that the yield by weight (weight of harvested mushrooms per 100 kg of compost) over 5 weeks of harvest is at least equal to 10%, and that the mushrooms produced have a normal appearance (morphology, dimensions).
The usual culture substrates are well known and are described, for example, in the abovementioned works. They are generally composts based on horse manure and straw.
The invention relates in particular to a hybrid mycelium as described hereinabove which is capable of forming fruiting bodies at a substrate temperature of 28xc2x0 C. or above and/or which is capable of forming fruiting bodies at an ambient temperature of 26xc2x0 C. or above, in particular at 28xc2x0 C. or above, and/or which is capable of multiplying on a usual culture substrate and of penetrating this substrate at a substrate temperature of 32xc2x0 C. or above without losing the capability of forming fruiting bodies, and/or which is capable of forming fruiting bodies without casing with a sporophore yield of at least 10 kg per 100 kg of culture substrate over a period of 5 weeks, starting at the beginning of the formation of fruiting bodies.
The hybrid mycelium according to the invention is generally heterokaryotic, but may also originate from a heterokaryotic or even homokaryotic single-spore culture of such a hybrid. It has indeed been discovered that, surprisingly, certain homokaryotic (that is to say haploid) mycelia which originate from hybrids according to the invention were capable of forming fruiting bodies; see experimental part hereinbelow.
The invention covers the sporophores which originate from the formation of fruiting bodies of a hybrid mycelium as defined hereinabove, as well as heterokaryons or homokaryons which originate from such a mycelium or such a sporophore.
The invention also relates to a method of growing Agaricus bisporus, which comprises a step in which mycelium is multiplied on a usual culture substrate and penetrates this substrate, followed by a step in which fruiting bodies are formed, in which this mycelium is a hybrid mycelium as defined above and exhibiting at least one of the following traits:
the process does not comprise a casing step;
the step in which the mycelium is multiplied on the substrate and the substrate is penetrated is carried out at a substrate temperature of 26xc2x0 C. or above;
the step in which fruiting bodies are formed is carried out at an ambient temperature of 24xc2x0 C. or above.
In particular embodiments, the production process according to the invention can also exhibit the following features, either alone or in combination:
the step in which the mycelium is multiplied and the substrate is penetrated is carried out at a substrate temperature of 28xc2x0 C. or above, in particular at 30xc2x0 C. or above;
the step in which fruiting bodies are formed is carried out at a substrate temperature of 28xc2x0 C. or above;
the step in which fruiting bodies are formed is carried out at an ambient temperature of 26xc2x0 C. or above, in particular at 28xc2x0 C. or above;
the process is carried out without the step of a thermal shock after the step in which the mycelium is multiplied.
The invention also relates to a method of obtaining a hybrid mycelium, sporophores or hetero-karyons or homokaryons as defined hereinabove, the process being characterized in that it comprises at least one hybridization step between a strain of Agaricus bisporus or a hybrid progeny of this strain and an Agaricus bisporus var. burnettii or a hybrid progeny of this variety, and in that those hybridization products are selected which exhibit the desired phenotypic trait. It follows from this that these phenotypic traits are genetically transmissible and are expressed in a stable fashion after having been transmitted.
The desired phenotypic character can be either one of the phenotypical traits mentioned in the definition of the hybrid mycelium according to the invention or a combination of two or more of these traits.
The variety burnettii of the species Agaricus bisporus has been described by Callac et al., Mycologia 85 (5), 835-851 (1993).
The hybridization steps can be carried out in a manner known per se. They are facilitated by the fact that, when Agaricus bisporus var. burnettii is hybridized with any strain of Agaricus bisporus, they produce hybrids whose spores are mostly homokaryotic; see, in this context, document WO 94/07357 and the U.S. Pat. No. 5304721.
To carry out the hybridization step in order to obtain what has been described, the Agaricus bisporus var. burnettii which may be used is, in particular, strain JB3 ms, which has been deposited at CNCM (France) on Jan. 23, 1997, under the number I-1811, and strain JB162 ms, which has been deposited at CNCM on Jan. 23, 1997, under the number I-1812.
An Agaricus bisporus which may be used as the first Agaricus bisporus (as defined in the process of obtaining it which has been described hereinabove) is, for example, one which does not belong to the variety burnettii, or a hybrid progeny of such an Agaricus bisporus. In particular, different strains of commercially obtainable mushrooms can be used, such as, for example, HU1. The strain HU1, which is also termed U1, is a commercial strain obtained from Station de Horst (Netherlands) and distributed by Sylvan-Somycel. This first Agaricus bisporus can also belong to the variety burnettii or be a hybrid progeny of this variety.
To carry out hybridization strategies, it is possible to use, in particular, the methods described in the PCT document WO 94/07357, which corresponds, in particular, to the U.S. Pat. application Ser. No. 08/403750, whose description is incorporated in the present description by way of reference.